In histopathology and cytopathology a pathologist routinely analyses microscopic images taken from tissues or cell smears. The specimens are typically analysed with a standard microscope using visible light. Due to the fact that cells and tissues hardly absorb visible light it is a common practice to stain the specimens. The staining chemical can be observed in visible light. It indicates the presence and generally also the amount of the structure it typically binds to. During many years of training and experience a pathologist learns how to interpret the stained images of specimens and to come to a diagnosis. In the digital pathology community the use of staining chemicals is generally considered mandatory for imaging slide samples in transmission.
Which staining method is chosen typically depends on the specific structure the pathologist is interesting in. A very popular staining method is the hematoxylin and eosin (H&E) stain. The hematoxylin binds to basophilic structures and colours them with a blue-purple hue. Basophilic structures are, for example, those cell components which contain nucleic acids and the cytoplasmic regions rich in RNA. Eosinophilic structures are generally composed of intracellular or extracellular protein and are coloured pink by the eosin.
However, staining has several major disadvantages. One of them is that the colour rendering in image of a stained specimen strongly depends on the stain method that has been used. The quality of the staining is not constant, as pointed out by T. Abe et al (T. Abe et al, “Colour Correction of Pathological Images Based on Dye Amount Quantification”. Optical Review, Vol. 12, No. 4, 2005, pp. 293-300). Yet a good quality is important for enabling the pathologist to come to a proper diagnosis. Another problem is related to the transfer of traditional pathology to automated diagnosis, which is an important development today. Quality variations observed among hospitals or even among staining machines or in staining machines over time present a serious hurdle for automated diagnosis. Different solutions have been proposed in order to solve these problems, such as improving the control of the staining process or by digitally correcting the multicolour image of the stained sample by means of a computer (see the above-mentioned article by T. Abe et al).
It is an object of the invention to provide a way of generating a multicolour image of an unstained biological specimen which does not require staining the specimen.
This object is achieved by the features of the independent claims. Further specifications and preferred embodiments are outlined in the dependent claims.